Nitroxidative Signaling Mechanisms in Pathological Pain

Abstract

Tissue injury can initiate bidirectional signaling between neurons, glia, and immune cells that creates and amplifies pain. While the ability for neurotransmitters, neuropeptides, and cytokines to initiate and maintain pain has been extensively studied, recent work has identified a key role for reactive oxygen and nitrogen species (ROS/RNS; nitroxidative species), including superoxide, peroxynitrite, and hydrogen peroxide. In this review we describe how nitroxidative species are generated after tissue injury and the mechanisms by which they enhance neuroexcitability in pain pathways. Finally, we discuss potential therapeutic strategies for normalizing nitroxidative signaling, which may also enhance opioid analgesia, to help to alleviate the enormous burden of pathological pain.

Keywords: NADPH oxidase; TRP channels; exercise; mitochondria; neuroinflammation; sensitization.

Figure 1. Induction of nitroxidative species after tissue injury

Nitroxidative species can induce posttranslational modifications of proteins and lipids, which subsequently drive pathological pain by modulating nociceptive neurotransmission, activating TRP channels, inducing mitochondrial dysfunction, and induce inflammatory signaling. In healthy cells, endogenous antioxidant systems prevent nitroxidative damage. Cell damage/pathology can perturb this balance, driving accumulation of potentially damaging nitroxidative species. O₂: oxygen; NO: nitric oxide; O₂^•−: superoxide; ONOO⁻: peroxynitrite; H₂O₂: hydrogen peroxide; ^•OH: hydroxyl radical; H₂O: water; NOX: NADPH oxidase; NOS: nitric oxide synathse; mETC: mitochondrial electron transport chain; SOD: superoxide dismutase; CAT: catalase; GPx: glutathione; HO: heme oxygenase.

Figure 2. Sources of nitroxidative species after tissue injury

Principal sources of nitroxidative species include NADPH oxidase (NOX), nitric oxide synthase (NOS), and electron leakage from the mitochondrial electron transport chain (mETC). The NOX1, 2, and 4 isoforms are differentially expressed across cell types and tissues after injury. NOX1-derived reactive oxygen species induce enhance Transient Receptor Potential (TRP) V1 activity in dorsal root ganglia (DRG) neurons. NOX2 activity in macrophages and microglia drives mRNA expression of proinflammatory cytokines (PIC) in DRG the spinal dorsal horn. NOX4 expression at the site of peripheral nerve injury decreases expression of myelin proteins (MP). The three NOS isoforms—NOS1 (neuronal), 2 (inducible), and 3 (endothelial)—are also differentially expressed by cell type. In abnormal pain states, N-methyl-D-aspartate receptors (NMDARs) are activated, resulting in calcium influx and activation of NOS1. Transcription of NOS2 is initiated by Toll like receptors (TLRs). These enzymes and processes have a well-established role in pathological pain.

Figure 3. Nitroxidative mechanisms of neuroexcitability after tissue injury

Reactive nitroxidative species, such as hydrogen peroxide and peroxynitrite, and modified proteins and lipids, like carbonylated proteins, peroxidated and nitrated lipids, and reactive aldehydes, all contribute to peripheral and central sensitization after tissue injury. These processes drive pathological pain. Several of the Transient Receptor Potential (TRP) family of nonselective cation channels are activated by nitroxidative species and modified proteins and lipids (see Nitroxidative species activate TRP channels). TRPA1 is expressed by peptidergic C-fibers, and is activated by modified proteins and lipids. TRPM2, which is expressed by neurons, monocytes/macrophages, microglia, and T cells, is directly activated by nitroxidative species. TRPM2 also activates intracellular signaling pathways, including mitogen activated protein kinase (MAPK) and nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells (NFκB) pathways. TRPV1 is found on C-fibers and is directly activated by some modified proteins and lipids, as well as being a target of oxidation and nitration events by nitroxidative species that increase responsiveness of the channel. Reactive nitroxidative species can directly modulate neuroexcitibility in central synapses by promoting glutamate release from primary afferent terminals, by activating calcium calmodulin-dependent protein kinase II (CamKII) in glutamatergic spinal neurons, and by inhibiting GABAergic interneurons (see Nitroxidative species as neuromodulators in pain pathways). Nitroxidative species also disrupt glutamate homeostasis by nitration and phosphorylation of NMDA receptor (NMDAR) subunits, as well as inhibiting glutamine synthetase (GS) and the glutamate transporter GLT-1. Mitochondrial DNA is a target of oxidation and nitration, while some nitroxidative species can form adducts with many mitochondrial proteins, which together impairs the structural integrity and function of mitochondria (see Nitroxidative species induce mitochondrial dysfunction). Nitroxidative species can also trigger release of pro-apoptotic factors from mitochondria by disrupting organelle dynamics. Nitroxidative species induce production of proinflammatory mediators, and can activate NFκB and MAPK intracellular signaling pathways (see Nitroxidative species induce neuroinflammatory signaling). Toll like receptors (TLRs) bind a variety of endogenous danger signals, including those released from nitroxidative-damaged mitochondria, to activate NFκB and MAPKs. NOX-derived ROS are second messengers for NFκB- and p38 MAPK-dependent TLR signaling, and TLR expression. The TLR2-NOX1 interaction also upregulates adhesion molecules via CCL3, which facilitates transendothelial cell migration into the CNS. Mitochondria-derived ROS also activate NLRP3 inflammasomes, which are protein complexes responsible for the proteolytic activation of IL-1β.